FIGURE 6.

Effects of H₂O₂ on transient depolarization of mitochondria in H9c2 cells. (A–C) TMRE fluorescence in H9c2 cells upon sequential addition of H₂O₂ and oligomycin. H₂O₂ (1 mM) and oligomycin (5 μM) were added in order. (A) Fluorescence images of TMRE in H9c2 cells. (B) The time course of changes in Cell TMRE averaged over 10 H9c2 cells. Arrows indicate the addition of H₂O₂ or oligomycin. The fluorescence at t = 0 was normalized to 100%. (C) Cell TMRE before the addition of H₂O₂ and oligomycin was normalized to 100%. When H₂O₂ or oligomycin was present, cells were incubated with H₂O₂ or oligomycin for 20 min before measurements. (D–F) TMRE fluorescence in H9c2 cells upon sequential addition of oligomycin and H₂O₂. Oligomycin (5 μM) and H₂O₂ (1 mM) were added in order. (D) Fluorescence images of TMRE in H9c2 cells. (E) The time course of changes in Cell TMRE averaged over 10 H9c2 cells. Arrows indicate the addition of H₂O₂ or oligomycin. The fluorescence at t = 0 was normalized to 100%. (F) Cell TMRE before addition of H₂O₂ and oligomycin was normalized to 100%. When H₂O₂ or oligomycin was present, cells were incubated with H₂O₂ or oligomycin for 20 min before measurements. (G) Frequency of transient depolarization of mitochondria in H9c2 cells. The frequency is shown as the number of depolarizations/min/cells. CCCP and CsA were added as described in figure legends for Figure 3. When H₂O₂ was present, cells were incubated with H₂O₂ for 20 min before measurements. (H) Statistical analysis of Cell TMRE in H9c2 cells. H₂O₂, CCCP and CsA were added as described for (G). Values represent the mean ± SEM (n > 10). *p < 0.05 vs. control for (G); without CCCP, H₂O₂ and CsA for (H).